大孔树脂吸附–交联法固定脂肪酶  

Immobilization of lipase on macroporous resin by adsorption-crosslinking method

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作  者:钱明华 张继福[2] 张云[3] 孙爱君[3] 于荣敏[1] 朱建华[1] 胡云峰[3] QIAN Minghua;ZHANG Jifu;ZHANG Yun;SUN Aijun;YU Rongmin;ZHU Jianhua;HU Yunfeng(Biotechnological Institute of Chinese Materia Medica, Ji'an University, Guangzhou 510632, China;Guangdong Provincial Hospital of Chinese Medicine, Guangzhou 510120, China;Chinese Academy of Sciences Key Laboratory of Tropical Marine Bio-resources and Ecology, SouthChina Sea Institute of Oceanology,Cliinese Academy of Sciences/Guangdong Key Laboratory of Marine Materia Medica, Guangzhou 510301, China)

机构地区:[1]暨南大学中药生物技术研究所,广东广州510632 [2]广东省中医院,广东广州510120 [3]中国科学院南海海洋研究所中国科学院热带海洋生物资源与生态重点实验室/广东省海洋药物重点实验室,广东广州510301

出  处:《华南农业大学学报》2019年第2期103-110,共8页JOURNAL OF SOUTH CHINA AGRICULTURAL UNIVERSITY

基  金:中国科学院战略性先导科技专项(XDA11030404);广东省海洋渔业科技攻关与研发方向项目(A201701C12);广州市科技计划项目(201510010012).

摘  要:【目的】为基于大孔树脂吸附结合环氧交联剂交联法固定脂肪酶等工业用酶奠定基础。【方法】使用大孔树脂吸附,而后环氧交联剂交联的方法进行脂肪酶的固定化,研究各因素对吸附–交联固定化的影响,并采用响应面法对固定化条件进行优化,制备固定化酶并考察其稳定性。【结果】筛选出大孔树脂HPD750为载体,聚乙二醇二缩水甘油醚为交联剂。最佳固定化条件为:吸附温度45℃,给酶量60 mg·g^–1,交联温度30℃,交联时间12.5 h,pH6.36,交联剂体积分数为0.7%。由上述条件制备所得的固定化酶活力为565.31 U·g^–1,酶活力回收率为32.16%。与游离酶相比,固定化酶的热稳定性和酸碱稳定性均有明显提升;连续操作10次,固定化酶活力仍保留34.86%,操作稳定性较好;4℃条件下储存30 d,固定化酶活力仍保留64.81%。【结论】大孔树脂HPD750为载体,聚乙二醇二缩水甘油醚为交联剂制备的固定化脂肪酶热稳定性、酸碱稳定性均得到显著提升,且具有良好的操作及储存稳定性。【Objective】To provide a basis for immobilization of industrial enzymes such as lipase using macroporous resin for adsorption and epoxy crosslinker for crosslinking.【Method】The immobilization of lipase was performed through adsorption using macroporous resin as the carrier and epoxy crosslinker as the crosslinking agent.We investigated the effects of different factors on adsorption-crosslinking immobilization,and used response surface design to optimize the immobilization conditions.The immobilized enzyme was prepared and its stability was investigated.【Result】Macroporous resin HPD750 was selected as the carrier,and poly(ethylene glycol)diglycidyl ether was selected as the crosslinker.The optimal conditions of lipase immobilization were as follows:Adsorption temperature 45°C,enzyme addition amount 60 mg·g^–1,crosslinking temperature 30 crosslinking time 12.5 h,pH6.36,and comcentration of crosslinking agent 0.7%.Under these optimized conditions,the immobilized lipase activity was 565.31 U·g^–1 and the recycled rate of lipase activity was 32.16%.Compared with free lipase,the immobilized lipase exhibited obviously better thermal stability and pH stability.The immobilized lipase was of good operation stability and remained 34.86%of the original activity after repeated usage for 10 times.The immobilized lipase also exhibited good storage stability and remained 64.81%of the original activity after storage at 4°C for 30 days.【Conclusion】Using macroporous resin HPD750 as the carrier and poly(ethylene glycol)diglycidyl ether as the crosslinker,the immobilized enzyme has significantly improved thermal stability and pH stability,and it also exhibits good operation stability and storage stability.

关 键 词:脂肪酶 大孔树脂 环氧交联剂 聚乙二醇二缩水甘油醚 固定 响应面法 

分 类 号:Q814.2[生物学—生物工程]

 

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